WO2012049228A2 - Marker sequences for multiple sclerosis and the use thereof - Google Patents

Abstract

The invention relates to novel marker sequences for multiple sclerosis and to the use thereof in diagnosis as well as to a method for screening potential active ingredients for multiple sclerosis diseases using said marker sequences. The invention further relates to a diagnostic device containing such marker sequences for multiple sclerosis, especially to a protein biochip and the use thereof.

Description

Marker sequences for multiple sclerosis and their use

description

The present invention relates to novel marker sequences for multiple sclerosis and their diagnostic use, including a method for screening potential drugs for multiple sclerosis disorders by means thereof

Marker sequences. Furthermore, the invention relates to a

diagnostic device containing such marker sequences for multiple sclerosis, in particular a protein biochip and its use.

Protein biochips are gaining an increasing industrial

Meaning in analytics and diagnostics as well as in the

Pharmaceutical development. Protein biochips have proven to be

Established screening tools.

Here, the fast and highly parallel detection of a variety of specific binding analysis molecules in one

single experiment allows. For production of

Protein biochips require the necessary proteins to be available. In particular

Protein expression libraries established. High throughput z cloning of defined open reading frames is one

Possibility (Heyman, JA, Cornthwaite, J., Foncerrada, L., Gilmore, JR, Gontang, E., Hartman, KJ, Hernandez, CL, Hood, R., Hull, HM, Lee, WY, Marcil, R. , Marsh, EJ, Mudd, KM, Patino, MJ, Purcell, TJ, Rowland, JJ,

Sindici, M.L. and Hoeffler, J.P. (1999) Genome-scale cloning and expression of individual open reading frames using

topoisomerase I-mediated ligation. Genome Res, 9, 383-392;

Kersten, B., Feilner, T., Kramer, A., Wehrmeyer, S., Possling, A., Witt, I., Zanor, MI, Stracke, R., Lueking, A., Kreut Zberger, J., Lehrach, H. and Cahill, DJ (2003)

Generation of Arabidopsis protein chip for antibody and serum screening. Plant Molecular Biology, 52, 999-1010; Reboul, J., Vaglio, P., Rual, J. F., Lamesch, P., Martinez, M., Armstrong,

C.M., Li, S., Jacotot, L., Bertin, N. , Janky, R., Moore, T., Hudson, J.R., Jr. Hartley, J.L., Brasch, M.A., Vandenhaute, J., Boulton, S., Endress, G.A., Jenna, S., Chevet, E.,

Papasotiropoulos, V., Tolias, P.P., Ptacek, J., Snyder, M., Huang, R., Chance, M.R., Lee, H., Doucette strain, L., Hill,

D.E. and Vidal, M. (2003) C. elegans ORFeome ersion 1.1:

experimental verification of the genome annotation and

resource for proteome-scale protein expression. Nat Genet, 34, 35-41; Walhout, A.J., Temple, G.F., Brasch, M.A., Hartley, J.L., Lorson, M.A., van den Heuvel, S. and Vidal, M. (2000)

GATEWAY recombinational cloning: application to the cloning of large numbers of open reading frames or ORFeems. Methods Enzymol, 328, 575-592). However, such an approach is strongly related to the progress of genome sequencing projects and the annotation of these gene sequences. In addition, the determination of the expressed sequence is due

differential splicing is not always clear. This problem can be overcome by the use of cDNA

Büssow, K., Cahill, D., Nietfeld, W., Bancroft, D., Scherzinger, E., Lehrach, H. and Walter, G. (1998) A method for global protein expression and antibody screening Nucleic Acids Research, 26, 5007-5008, Büssow, K., Nordhoff, E., Lübbert, C, Lehrach, H. and Walter, G. "

(2000) A human cDNA library for high-throughput protein expression screening. Genomics, 65, 1-8; Holz, C, Lueking, A., Bovekamp, L., Gutjahr, C, Bolotina, N., Lehrach, H. and Cahill, DJ (2001) A human cDNA expression library in yeast enriched for open reading frames. Genome Res, 11, 1730-1735; Lueking, A., Holz, C, Gotthold, C, Lehrach, H. and Cahill, D. (2000) A system for dual protein expression in Pichia pastoris and Escherichia coli, protein Expr. Purif. , 20, 372-378). Here, the cDNA of a particular tissue in a bacterial or a eukaryotic expression vector, such as yeast, is cloned. The vectors used for the expression are generally characterized by the fact that they carry inducible promoters, with which the timing of protein expression can be controlled. In addition, expression vectors have sequences for so-called

Affinity epitopes or proteins, on the one hand for the specific detection of the recombinant fusion proteins by means of a directed against the affinity epitope

 Antibody, on the other hand becomes the specific one

Purification via affinity chromatography (IMAC) allows.

For example, the gene products of a cDNA expression library of human fetal brain tissue in the bacterial expression system Escherichia coli in high density format were placed on a membrane and successfully screened with different antibodies. It could be shown that the proportion of full-length proteins is at least 66%. The recombinant proteins out

In addition, expression libraries have been published in

 High throughput can be expressed and purified (Braun P., Hu, Y., Shen, B., Halleck, A., Koundinya, M., Harlow, E. and LaBaer, J. (2002) Proteome-scale purification of human

proteins from bacteria. Proc Natl Acad. USA, 99, 2654-2659; Büssow (2000) supra; Lueking, A., Horn, M., Eickhoff, H., Büssow, K., Lehrach, H. and Walter, G. (1999) Protein microarrays for gene expression and antibody screening.

Analytical Biochemistry, 21Q, 103-111). Such protein biochips are based on cDNA expression libraries

in particular the subject matter of WO 99/57311 and WO 99/57312. Further, in addition to antigen-presenting protein biochips, antibody-presenting arrangements are also described (Lal et al (2002) Antibody arrays: An embryonic but growing technology, DDT, 7, 143-149, Kusnezow et al., (2003) Antibody microarrays: An evaluation of production parameters, Proteomics, 3, 254-264).

However, there is a great need for indication-specific diagnostic devices, such as a protein biochip,

to provide.

The object of the present invention is to provide improved marker sequences and their diagnostic

Use for the treatment of multiple sclerosis.

The provision of specific marker sequences allows a reliable diagnosis and stratification of patients with multiple sclerosis, in particular by means of a protein biochip. Therefore, the invention relates to the use of

 Marker sequences for the diagnosis of multiple sclerosis, wherein at least one marker sequence of a cDNA selected from the group SEQ 1-81 or in each case a protein coding therefor or in each case a partial sequence or fragment thereof

(hereinafter: marker sequences according to the invention) to or from a patient to be examined.

The marker sequences of the invention could by means of

differential screening of samples, and more healthy Subjects with patient samples identified with multiple sclerosis.

For the first time protein biochips (see

Examples) of these marker sequences according to the invention

be identified.

Although in the prior art marker sequences for multiple sclerosis could already be identified with the aid of a protein biochip, see WO2009030225. In the present case however

According to the invention, an improved bioinformatic evaluation is strained and the samples are particularly preferably taken from the cerebrospinal fluid (CSF). In addition, specially selected samples are used that meet the high sensitivity of a protein biochip.

The term "multiple sclerosis ((MS), also encephalomyelitis disseminata)" refers to an autoimmune inflammatory /

demyelinating and degenerative disease of central nervous system disease (definition, for example, according to Pschyrembel, de Gruyter, 261st edition (2007), Berlin).

Essential to the invention is that the samples are not taken from conventional blood banks, but from MS patients

have been carefully selected, for example, HIV and HCV are negative and in particular have been tested for infectious diseases. The complex sample selection method allows, for example, a sufficient advantageous delimitation of diseases such as MS symptom-like neuroborelliosis. Furthermore, false-positive results are excluded, on the one hand due to the strict bioinformatory evaluation (see examples) and by comparing the results on a protein biochip according to the invention with eg sera from Neuroborelliosis patients who do not have multiple sclerosis

exhibit .

Furthermore, in contrast to WO2009030225, the

Production of the protein biochips by normalization of at least 1,000, preferably 2,000 different or more autoantigens of humans, which are not indication-specific for multiple sclerosis. Such autoantigens may e.g. from other body fluids of patients of others

Diseases (e.g., pancreatic cancer, rheumatoid arthritis, prostate, etc.).

The invention therefore also relates to such indication-specific protein biochips according to the invention for the diagnosis of

Multiple sclerosis, whereby in a further step the proteins or proteins represented on the protein biochip

Marker sequences with autoantibodies from non-multiple

Sclerosis patients can be normalized and in this way false-positive proteins can be removed. Remaining non-false positive proteins can be reassembled on a protein biochip, called rearraying. This also allows the exclusion of autoantibodies that are positive for E. coli. This is another qualitative

Improvement, since autoantibodies can be excluded, e.g. directed against E. coli intestinal bacteria in humans. As a result, new marker sequences can be advantageously identified with an improved signal-to-noise ratio.

In a further preferred embodiment, therefore, at least 2 to 5 or 10, preferably 30 to 50

Marker sequences or 50 to 81 or more marker sequences to or from a patient to be examined. In a further embodiment of the invention, the marker sequences according to the invention can also be combined, supplemented or extended with known biomarkers for this indication. In a preferred embodiment, the determination of the marker sequences takes place outside the human body and the determination is made in an ex vivo / in vitro diagnosis.

In a further embodiment of the invention, the invention relates to the use of marker sequences as diagnostics, wherein at least one marker sequence of a cDNA is selected from the group SEQ 1-81 or in each case a protein coding therefor or in each case a partial sequence or fragment thereof.

Furthermore, the invention relates to a method for the diagnosis of multiple sclerosis, wherein a.) At least one marker sequence of a cDNA selected from the group SEQ 1-81 or a respective coding protein or a partial sequence or fragment thereof is applied to a solid support and b .) with body fluid or tissue extract one

C.) The detection of an interaction of the body fluid or tissue extract with the marker sequences from a.) Is done.

Therefore, the invention also relates to diagnostics for

Diagnosis of multiple sclerosis in each case selected from the group SEQ 1-81 or in each case a protein coding therefor or in each case a partial sequence or fragment thereof.

The detection of such an interaction can, for example, by a probe, in particular by an antibody

respectively . Therefore, the invention also relates to the task of providing a diagnostic device or an assay, in particular a protein biochip, which allows a diagnosis or examination for multiple sclerosis. Furthermore, the invention relates to a method for

 Stratification, in particular for the risk stratification and / or therapy control of a patient with multiple sclerosis, wherein at least one marker sequence of a cDNA selected from the group SEQ 1-81 or a protein coding therefor is determined on a patient to be examined.

Also included is the stratification of patients with

Multiple sclerosis into new or established subgroups of

Multiple sclerosis, as well as the sensible selection of

Patient groups for the clinical development of new ones

Therapeutics. The term therapy control also includes the classification of patients into responders and non-responders with regard to a therapy or its course of therapy.

"Diagnosis" in the sense of this invention means the positive finding of multiple sclerosis by means of

Marker sequences of the invention and the assignment of patients to the disease of multiple sclerosis. The term diagnosis includes medical diagnostics and

related studies, in particular in vitro diagnostics and laboratory diagnostics, also proteomics and

Nucleic acid blots. Further investigations can be made for

Protection and exclusion of other diseases. Therefore, the term diagnosis also includes the

Differential diagnosis of multiple sclerosis using the

Marker sequences according to the invention and the prognosis of

Multiple sclerosis. "Stratification (also: stratification) or therapy control" in the sense of this invention means that the inventive method decisions for the treatment and therapy of the

Patients, whether it be hospitalization of the patient, use, effect and / or dosage of one or more drugs, a therapeutic measure or the

Surveillance of disease progression as well as treatment course or etiology or classification of a disease, e.g. into a new or existing subtype or the differentiation of diseases and their patients.

In a further embodiment of the invention, the term "stratification" includes in particular the

 Risk stratification with the prognosis of an "outcome" of a detrimental health event For the purposes of this invention, "patient" means any subject - human or mammal - with the proviso that the subject is being examined for multiple sclerosis.

The term "marker sequences" in the sense of this invention means that the cDNA or the respective polypeptide or protein obtainable therefrom are significant for multiple sclerosis For example, the cDNA or the respectively obtainable polypeptide or protein can interact with substances from the body fluid or tissue extract of a Patients with multiple sclerosis (eg antigen

(Epitope) / antibody (paratope) interaction). For the purposes of the invention, "at least one marker sequence of a cDNA selected from the group SEQ 1-81 or in each case a protein coding therefor or in each case a partial sequence or fragment thereof to or from a patient to be examined is determined" that an interaction between the Body fluid or tissue extract of a patient and the marker sequences of the invention is detected. Such an interaction is for example a bond, in particular a binding substance on at least one invention

Marker sequence or, in the case of a cDNA, hybridization with a suitable substance under selected conditions, in particular stringent conditions (for example, as usual

defined in J. Sambrook, E.F. Fritsch, T. Maniatis (1989), Molecular cloning: A laboratory manual, 2nd Edition, Cold Spring Habor Laboratory Press, Cold Spring Habor, USA, or Ausubel, "Current Protocols in Molecular Biology," Green

Publishing Associates and Wiley Interscience, N.Y. (1989)). An example of stringent hybridization conditions is: hybridization in 4 × SSC at 65 ° C. (alternatively in 50%

Formamide and 4X SSC at 42 ° C), followed by several

 Washes in 0.1 x SSC at 65 ° C for a total of about one hour. An example of less stringent

 Hybridization conditions are hybridization in 4 x SSC at 37 ° C followed by several washes in 1 x SSC at room temperature.

Such substances are part of a component according to the invention

Body fluid, in particular blood, whole blood, blood plasma, blood serum, patient serum, urine, cerebrospinal fluid, synovial fluid or a tissue extract of the patient. In a further embodiment of the invention, however, the marker sequences according to the invention may be present in a significantly higher or lower expression rate or concentration, which points to multiple sclerosis. In this case, by means of proteomics or nucleic acid blots, the relative expression rates ill / healthy of the invention

Marker sequences for multiple sclerosis are determined. In a further embodiment of the invention, the marker sequences have a recognition signal which is addressed to the substance to be bound (for example antibodies,

Nucleic acid). According to the invention, for a protein, the recognition signal is preferably an epitope and / or paratope and / or hapten and, for a cDNA, a hybridization or hybridization protein

Binding region.

The marker sequences of the invention are the subject of Table A and can by the respective cited

Database entry (also via Internet:

http://www.ncbi.nlm.nih.gov/) are clearly identified (see Table A: Accession No. there), see also the corresponding sequence listing.

Therefore, the invention also relates to the full-length sequences of the markers according to the invention and indeed as defined in Table A on the known database entry, hereinafter called SEQ -a 81a (cDNA) or SEQ ID-81b (protein).

Furthermore, therefore, also include analog

 Embodiments of SEQ ID NO: 81a to the marker sequences SEQ ID-81, e.g. set forth in the claims, since the

SEQ 1-81 according to the invention again represent partial sequences, at least with high homology. The specific ones

However, marker sequences SEQ 1-81 are in accordance with the invention

prefers . According to the invention, the marker sequences also include such

Modifications of the cDNA sequence and the corresponding

Amino acid sequence, such as chemical modification, such as

Citrullination, acetylation, phosphorylation,

Glycation or polyA strand and other modifications known to those skilled in the art. In a further embodiment of the invention are also partial sequences or fragments of the invention

Includes marker sequences. In particular, those partial sequences which have an identity of 95%, 90%, in particular 80% or 70% with the marker sequences according to the invention.

Partial sequences are also those sequences having 50 to 81 nucleotides, 70-120 nucleotides of a sequence of SEQ 1-81, or peptides obtainable therefrom.

"Partial sequences or fragments" of the invention

Marker sequences are functionally defined and include those sequences which have the same inventive diagnostic function.

In a further embodiment, the respective

Marker sequence can be represented in different amounts in one or more areas on a solid support. This allows a variation of the sensitivity. The regions may each comprise a total of marker sequences, i. a sufficient number of different marker sequences, in particular 2 to 5 or 10 or more and optionally further nucleic acids and / or proteins, in particular biomarkers.

However, at least 96 to 25,000 (numerically) or more from different or identical marker sequences and further nucleic acids and / or proteins, in particular, are preferred

Biomarkers. Further preferred are more than 2,500, more preferably 10,000 or more different or the same

 Marker sequences and optionally other nucleic acids and / or proteins, in particular biomarkers.

Another object of the invention relates to an array of marker sequences containing at least one marker sequence a cDNA selected from the group SEQ 1-81 or in each case a protein coding therefor. Preferably, the

Arrangement of at least 2 to 5 or 10, preferably 30 to 50 marker sequences or 50 to 81 or more marker sequences.

In the context of this invention, "arrangement" synonymously means "array" and insofar as this "array" is used to identify substances on marker sequences, this is to be understood as meaning an "assay" or a diagnostic device. In a preferred embodiment, the arrangement is designed such that those represented on the assembly

Marker sequences are in the form of a grid on a solid support. Furthermore, such arrangements are preferred which include a high density array of protein binders

allow and the marker sequences are spotted. Such high density spotted assemblies are disclosed, for example, in WO 99/57311 and WO 99/57312, and may be advantageously used in a robotic automated high throughput method.

In the context of this invention, however, the term "assay" or diagnostic device also includes such

Embodiments of a device such as ELISA, bead-based assay, line assay, Western blot, immunochromatographic

Methods (e.g., so-called lateral flow immunoassays) or similar immunological single or multiplex detection methods. A protein biochip in the sense of this

Invention is the systematic arrangement of proteins on a solid support.

The marker sequences of the assembly are fixed to a solid support, but preferably spotted or immobilized imprinted, ie applied reproducible. One or more marker sequences can be present multiple times in the totality of all marker sequences and be present in different amounts relative to one spot. Furthermore, the marker sequences can be standardized on the solid support (eg by serial dilution series of eg

Human globulins as internal calibrators for

Data normalization and quantitative evaluation).

Therefore, the invention relates to an assay or protein biochip consisting of an array containing marker sequences according to the invention.

In a further embodiment, the marker sequences are present as clones. Such clones can be obtained, for example, by means of a cDNA expression library according to the invention (Büssow et al., 1998 (supra)). In a preferred embodiment, such expression libraries become

containing clones by means of expression vectors from a

expressing cDNA library consisting of the cDNA

Obtained marker sequences. These expression vectors preferably contain inducible promoters. The induction of

 Expression can e.g. by means of an inductor, such as IPTG. Suitable expression vectors are described in Terpe et al. (Terpe T Appl Microbiol Biotechnol 2003 Jan; 60 (5): 523-33). Expression libraries are known to the person skilled in the art, these can be prepared according to standard works, such as Sambrook et al., Molecular Cloning, Laboratory Handbook, 2nd edition (1989), CSH press, Cold Spring Harbor, New York Further preferred are such expression libraries

are tissue specific (eg human tissue, in particular human organs). Furthermore, such expression libraries are also included according to the invention, which can be obtained by exon trapping. Instead of expression library can be spoken synonymously from an expression bank. Also preferred are protein biochips or corresponding expression libraries which have no redundancy (so-called: Uniclone® library) and can be prepared, for example, according to the teachings of WO 99/57311 and WO 99/57312. These preferred Uniclone libraries have a high content of non-defective, fully expressed proteins of a cDNA expression library.

Also within the scope of this invention, the clones may not be such as transformed bacteria, recombinant phage or transformed cells of mammals, insects, fungi, yeasts or plants.

The clones are fixed on a solid support, spotted or immobilized.

Therefore, the invention relates to an arrangement, wherein the

Marker sequences are present as clones. In addition, the marker sequences may be in the form of a fusion protein in the particular form

For example, at least one affinity epipope or "tag" contains. The tag may be one such as c-myc, His-tag, Arg-tag, FLAG, alkaline phosphatase, V5 tag, T7 tag or Strep tag, HAT tag, NusA, S-tag, SBP tag , Thioredoxin, DsbA, a fusion protein, preferably a cellulose-binding one

Domain, green fluorescent protein, maltose binding

Protein, calmodulin-binding protein, glutathione S-transferase or lacZ. In all embodiments, the term "solid support" includes embodiments such as a filter, a membrane, a magnetic or fluorophore-labeled bead

Silicon wafer, glass, metal, plastic, a chip, a mass spectrometric target or a matrix. However, a filter is preferred according to the invention.

Also preferred as the filter is PVDF, nitrocellulose or nylon (e.g., Immobilon P Millipore, Protran Whatman, Hybond N + Amersham). In a further preferred embodiment of the

According to the invention, this corresponds to a grid having the order of a microtiter plate (8-12 wells strips, 96 wells, 384 wells or more), a silicon wafer, a chip, a mass spectrometric target or a matrix.

In a further embodiment, the invention relates to an assay or protein biochip for identifying and

Characterizing a substance for multiple sclerosis, characterized in that an arrangement or assay according to the invention is brought into contact with a.) At least one substance to be investigated and b.) A binding success is detected.

Furthermore, the invention relates to a method for identifying and characterizing a substance for multiple sclerosis, characterized in that an arrangement or assay according to the invention is brought into contact with a.) At least one substance to be investigated and b.) A binding success

is detected. The substance to be tested may be any native or non-native biomolecule, a synthetic chemical

Be a molecule, a mixture or a substance library.

After the substance to be examined contacts a marker sequence, the evaluation of the binding success

performed, for example, using with

commercially available image analysis software (GenePix Pro (Axon Laboratories), Aida (Raytest), ScanArray (Packard Bioscience).) The visualization of protein-protein interactions of the invention (e.g., protein to marker sequence, such as

Antigen / antibody) or corresponding "means for detecting the binding success" can, for example, by means of

Fluorescent labeling, biotinization, radio-isotopic labeling or colloidal gold or latex particle marking in a conventional manner. Detection of bound antibodies takes place with the help of secondary antibodies

Antibodies with commercial reporter molecules

(e.g., Cy, Alexa, Dyomics, FITC or similar fluorescent dyes, colloidal gold or latex particles) or with reporter enzymes such as alkaline

Phosphatase, horseradish peroxidase, etc. and the

corresponding colorimetric, fluorescents or

chemoluminescent substrates. A readout is e.g. by means of a microarray laser scanner, a CCD camera or visually.

In a further embodiment, the invention relates to a drug / prodrug or prodrug developed for multiple sclerosis and obtainable through the use of the

Assays according to the invention or protein biochip. Therefore, the invention also relates to the use of an arrangement according to the invention or an assay for the screening of drugs for multiple sclerosis.

Therefore, in a further embodiment, the invention also relates to a target for the treatment and therapy of multiple sclerosis, each selected from the group SEQ 1-81 or in each case a protein coding therefor.

In a further embodiment, the invention also relates to the use of the invention

Marker sequences, preferably in the form of an arrangement, as

Affinity material for performing an apheresis or iwS. a blood wash, wherein substances from body fluids of a patient with multiple sclerosis, such as blood or plasma, bind to the marker sequences of the invention and thus the body fluid can be selectively withdrawn.

Examples and figures:

Ten or more patient samples were individually screened against a cDNA expression library. Multiple sclerosis - specific expression clones were identified by comparison with ten or more healthy specimens. The identity of the marker sequences was determined by DNA sequencing.

FIG. 1 shows the differential screening between two protein biochips from in each case one cDNA expression bank of a patient and one healthy subject. The

Differential clones are detected by fluorescence labeling and evaluated bioinformatorisch.

In the context of biomarker identification various bioinformatic analyzes are carried out. For each serum, microarray reactivities against about 2000 measured different antigens. These data are used for a ranking of the spotted antigens regarding their

Differentiation ability between healthy and diseased sera used. This evaluation is by means of not

parametric Mann-Whitney tests on normalized

 Intensity data performed. For normalization, an internal standard is used, which is spotted on each chip. Since a p-value is calculated for each antigen, methods for correcting the multiple testing are used. As a very conservative approach, a Bonferroni correction is performed and, in addition, the less restrictive False Discovery Rate (FDR) is calculated according to Benjamini & Hochberg.

Furthermore, the data are used to classify the sera. Here, different multivariate methods are used. These are methods from the statistical

 Learning methods such as support vector machines (SVM), neural networks or classification trees, as well as a

 Threshold method, which is suitable for both classification and visual representation of the data. To avoid overfitting, a 10-fold cross-validation of the data is performed.

Table A: (gi Accession number with validity from 1.10.2010)

SEQ ID NO: 1-81b SEQ ID NO: 1 a-81 a

gi Acc Protein gi Acc cDNA NAME

gi 149363636 plexin B2 [Homo sapiens]

gi 149363636 gi 149363635 plexin B2 [Homo sapiens]

gi 13259508 gi | 13259507 dynactin 1 isoform 2 [Homo sapiens]

gi 13259508 gi 13259507 dynactin 1 isoform 2 [Homo sapiens]

 DIS3 mitotic control homologue (S. cerevisiae) -like 2 gi | 148596939

gi1134288890 [Homo sapiens]

 transcription elongation factor B polypeptides 3 gi | 145199236

gi | 145199237 binding protein 1 [Homo sapiens] transcription elongation factor B polypeptides 3 gi | 145199236

gi 145199237 binding protein 1 [Homo sapiens]

gi 145309326 gi | 145309325 laminin, gamma 1 precursor [Homo sapiens] gi 145309326 gi 145309325 laminin, gamma 1 precursor [Homo sapiens] gi 157266266 gi | 157266265 WD repeat domain 86 [Homo sapiens]

gi 16975484 gi | 92091602 centaurin delta 2 isoform b [Homo sapiens] gi 16975484 gi 92091602 centaurin delta 2 isoform b [Homo sapiens] gi 20070228 gi | 39725676 nucleobindin 1 [Homo sapiens]

 hematological and neurological expressed 1-like gi | 46361989

gi | 21700763 [Homo sapiens]

gi 21707902 gi 21707901 CTTN protein [Homo sapiens]

 chromosome 20 open reading frame 3 [Homo gi | 41327713

gi | 24308201 sapiens]

gi 24797103 gi 24797102 RAS guanyl releasing protein 2 [Homo sapiens] gi 26051235 gi | 26051234 nucleoporin 133kDa [Homo sapiens]

gi 26051235 gi 26051234 nucleoporin 133kDa [Homo sapiens]

gi 29788785 gi | 34222261 tubulin, beta [Homo sapiens]

 F-box only protein, helicase, 18 isoform 2 [Homo gi | 307951 18

gi | 307951 19 sapiens]

 F-box only protein, helicase, 18 isoform 2 [Homo gi | 307951 18

gi | 307951 19 sapiens]

 SR-related CTD-associated factor 1 [Homo gi | 32698749

gi | 32698750 sapiens]

 SR-related CTD-associated factor 1 [Homo gi | 32698749

gi | 32698750 sapiens]

gi 33469964 gi | 33469963 splicing factor 4 [Homo sapiens]

 tubulin, gamma complex associated protein 4 gi | 38454193

gi | 38454194 [Homo sapiens]

 mannosidase, alpha, class 2A, member 2 [Homo gi | 51477715

gi | 51477716 sapiens]

 mannosidase, alpha, class 2A, member 2 [Homo gi | 51477715

gi | 51477716 sapiens]

gi 58331 179 gi 58331 178 KIAA1688 protein [Homo sapiens]

gi 58331 179 gi 58331 178 KIAA1688 protein [Homo sapiens]

gi 6005747 gi | 54792140 ring finger protein 2 [Homo sapiens]

gi 7706359 gi 22027484 RAS, dexamethasone-induced 1 [Homo sapiens] gi 1 12382377 gi | 1 12382376 ubiquitin-conjugating enzymes E2S [Homo sapiens] gi 4505677 gi 24431942 Phosphodiesterase 1 B [Homo sapiens]

gi 33636722 gi | 45827807 plasticity related genes 1 [Homo sapiens] gi 19526471 gi | 62739177 rhotekin isoform b [Homo sapiens]

gi 1 16812573 gi 148833501 CWC15 homolog [Homo sapiens]

 pyrroline-5-carboxylate reductase 1 isoform 2 gi | 24797094

gi | 24797095 [Homo sapiens]

gi 83035136 gi | 21362004 F-box protein 31 [Homo sapiens] zinc finger and BTB domain containing 5 [Homo gi | 7662073

gi | 7662074 sapiens]

 cyclin D binding myb-like transcription factor 1 gi | 48976050

gi | 48976051 [Homo sapiens]

gi 17986283 gi | 17986282 tubulin, alpha 1 a [Homo sapiens]

 splicing factor, arginine / serine-rich 10 [Homo gi | 215422394

gi | 4759098 sapiens]

 51477701 dependent regulator of chromatin, subfamily d, gi | 51477702 member 3 isoform 2 [Homo sapiens]

 IQ motif and WD repeats 1 isoform a [Homo gi | 63252907

gi | 63252908 sapiens]

 poliovirus receptor related 2 isoform alpha gi | 1 12789531

gi 11 12789532 precursor [Homo sapiens]

gi 22027541 g 22027540 programmed cell death 7 [Homo sapiens] gi 5902122 g 15902121 spectrin, beta, non-erythrocytic 2 [Homo sapiens] gi 5902122 g 5902121 spectrin, beta, non-erythrocytic 2 [Homo sapiens] gi 5902122 g 5902121 spectrin, non-erythrocytic 2 [Homo sapiens] gi 71361682 g 171361681 nuclear mitotic apparatus protein 1 [Homo sapiens] gi 71361682 g 71361681 nuclear mitotic apparatus protein 1 [Homo sapiens] gi 71361682 g 171361681 nuclear mitotic apparatus protein 1 [Homo sapiens] triple functional domain (PTPRF interacting) [Homo gi | 45439358

gi | 45439359 sapiens]

 triple functional domain (PTPRF interacting) [Homo gi | 45439358

gi | 45439359 sapiens]

 triple functional domain (PTPRF interacting) [Homo gi | 45439358

gi | 45439359 sapiens]

 triple functional domain (PTPRF interacting) [Homo gi | 45439358

gi | 45439359 sapiens]

 HECT, UBA and WWE domain containing 1 [Homo gi | 61676187

gi | 61676188 sapiens]

 HECT, UBA and WWE domain containing 1 [Homo gi | 61676187

gi | 61676188 sapiens]

 HECT, UBA and WWE domain containing 1 [Homo gi | 61676187

gi | 61676188 sapiens]

 HECT, UBA and WWE domain containing 1 [Homo gi | 61676187

gi | 61676188 sapiens]

 HECT, UBA and WWE domain containing 1 [Homo gi | 61676187

gi | 61676188 sapiens]

gi 95147333 gi 95147332 phospholipase C, beta 2 [Homo sapiens] gi 95147333 gi | 95147332 phospholipase C, beta 2 [Homo sapiens] gi 450481 1 g 213972609 junction plakoglobin [Homo sapiens]

 amyloid precursor-like protein 1 isoform 1 gi | 67782337

gi | 67782338 precursor [Homo sapiens]

gi 72534684 gi | 166197669 phospholipase D3 [Homo sapiens] gi 13128862 gi | 157266338 histone deacetylase 3 [Homo sapiens] gi 194018520 gi 194018519 G1 to S phase transition 1 [Homo sapiens]

 spectrin, beta, non-erythrocytic 1 isoform 1 [Homo gi | 1 12382249

gi | 1 12382250 sapiens]

 spectrin, beta, non-erythrocytic 1 isoform 1 [Homo gi | 1 12382249

gi 11 12382250 sapiens]

 spectrin, beta, non-erythrocytic 1 isoform 1 [Homo gi | 1 12382249

gi 11 12382250 sapiens]

gi 5031905 gi 187828416 MyoD family inhibitor [Homo sapiens]

gi 53759122 adenomatous polyposis coli [Homo sapiens] gi 53759122 gi 53759121 adenomatous polyposis coli gi 53759122 gi | 53759121 adenomatous polyposis coli [Homo sapiens] gi 40548332 gi 149363677 coiled-coil domain containing 137 [Homo sapiens]

 Wiskott-Aldrich Syndrome protein family member 1 gi | 68161503

gi | 68161504 [Homo sapiens]

 coenzyme Q10 homologous A isoform b [Homo gi | 151 101385

gi | 151 101386 sapiens]

gi 89903008 gi 237858673 Neurofascin

gi 89903008 gi | 237858674 Neurofascin

Claims

claims Use of marker sequences for the diagnosis of  Multiple sclerosis, wherein at least one marker sequence of a cDNA selected from the group SEQ 1-81 or in each case a protein coding therefor or in each case a partial sequence or fragment thereof to or from a patient to be examined is determined. 2. Use of marker sequences for the diagnosis of  Multiple sclerosis, wherein at least one marker sequence of a cDNA selected from the group SEQ. La - 81a or in each case a protein coding therefor or in each case a partial sequence or fragment thereof to or from a patient to be examined is determined. Use of marker sequences for the diagnosis of  Multiple sclerosis according to claim 1 or 2, wherein the sample is obtained from a patient to be examined from cerebrospinal fluid (CSF). 4. Use of marker sequences for the diagnosis of  Multiple sclerosis according to claim 1, 2 or 3, characterized in that at least 2 to 5 or 10, preferably 30 to 50 marker sequences or 50 to 81 or more marker sequences on or from one to  determined by the examining patient. Use of marker sequences for the diagnosis of Multiple sclerosis according to claim 1 or 2, characterized in that the determination is carried out by means of in vitro diagnosis. Use of a marker sequence of a cDNA in each case selected from the group SEQ 1-81 or SEQ.lcr.-81a or in each case a protein coding therefor or in each case a partial sequence or fragment thereof as a diagnostic agent. 7. Use of marker sequences for the diagnosis of  Multiple sclerosis after one of the previous ones  Claims, characterized in that the  Marker sequences are applied to a solid support, in particular a filter, a membrane, a magnetic or fluorophore-labeled bead, a silicon wafer, glass, metal, plastic, a chip, a mass spectrometric target or a matrix. 8. Method for the diagnosis of multiple sclerosis, wherein  a. ) at least one marker sequence of a cDNA selected from the group SEQ 1-81 or or SEQ. la 81a or a protein coding therefor or in each case a partial sequence or fragment thereof is applied to a solid support, and  b. ) with body fluid or tissue extract one  Patient is brought in contact and  c. ) the proof of an interaction of the  Body fluid or tissue extract with the  Marker sequences from a.) Takes place. Method for the diagnosis of multiple sclerosis according to  Claim 8, wherein in a further step  represented marker sequences, in particular proteins, with autoantibodies from non-multiple sclerosis  Patients are normalized. 10. Stratification method, in particular for Risk stratification, or therapy control of a Patients with multiple sclerosis, wherein at least one marker sequence of a cDNA selected from the group SEQ 1 - 81 or SEQ - 81a or one each for  coding protein or in each case a partial sequence or fragment thereof to or from a patient to be examined is determined. The method of claim 8, wherein the stratifying  or the therapy control decisions  Treatment and therapy of the patient, in particular hospitalization of the patient, use, effect and / or dosage of one or more drugs, a therapeutic measure or the monitoring of a disease course and therapy course, etiology or classification of a disease including prognosis. 12. Arrangement of marker sequences containing at least  a marker sequence of a cDNA selected from the group SEQ 1 - 81 or SEQ - 81a or in each case a protein coding therefor. 13. Arrangement according to claim 12, characterized in that at least 2 to 5 or 10, preferably 30 to 50 marker sequences or 50 to 81 or more  Marker sequences are included. 14. Arrangement according to claim 12, characterized in that the marker sequences are present as clones. 15. Assay, protein biochip consisting of an assembly according to claim 12, characterized in that the marker sequences are applied to a solid support. 16. Use of an arrangement according to one of claims 12 to 14 or an assay according to claim 15 for  Identification and characterization of a substance for multiple sclerosis comprising means for detecting a binding success, characterized in that an arrangement or assay is brought into contact with a.) At least one substance to be investigated and b.) A binding success is detected. 17. Use of an arrangement according to any one of claims 12 to 14 or an assay according to claim 15 for the screening of drugs for multiple sclerosis. 18. Diagnostics for the diagnosis of multiple sclerosis, each selected from the group SEQ 1-81 or SEQ.alpha.181a or in each case a protein coding therefor or in each case a partial sequence or fragment thereof. 19. Target for the treatment and therapy of multiple  Sclerosis, each selected from the group SEQ 1 - 81 or SEQ ID - 81a or in each case a protein coding therefor or in each case a partial sequence or fragment thereof. 20. Use of a marker sequence of a cDNA selected from the group SEQ 1-81 or SEQ.alpha.181a respectively a protein coding therefor or in each case one  Partial sequence or fragment thereof as affinity material for performing apheresis or blood washing for patients with multiple sclerosis.